Technical Abstract:
Riboflavin synthase (RS) catalyses the dismutation of two molecules of 6,7-dimethyl-8-(1´-D-ribityl)-lumazine (DMRL) to yield riboflavin and 4-ribitylamino-5-amino-2,6-dihydroxypyrimidine (RAADP). One four-carbon unit is transferred from the donor DMRL, which becomes RAADP, to the acceptor DMRL, which becomes riboflavin. RS is a homotrimer of 23 kDa subunits with each subunit consisting of two similar beta barrels and a C-terminal alpha helix. Active sites are formed at the interface between an N barrel (beta barrel belonging to the N terminus) of one subunit and a C barrel (beta barrel belonging to the C terminus) of another subunit. Only one active site of the three in the trimer can be formed at a time. We report the X-ray structure of the Escherichia coli RS in complex with riboflavin. Five riboflavins reside in the homotrimer, with one bound to each beta barrel except the C barrel that is part of the formed active site. The N barrel of the formed active site is occupied by riboflavin. This result substantiates models of the Michaelis complex and the pentacyclic reaction intermediate, which are based on the acceptor DMRL molecule belonging to the N barrel and the donor DMRL molecule belonging to the C barrel.